Help need for Single-step RNA Ioslation by Acid Guanidinium Thiocyanate-Phenol-Chloroform Extraction"

Traci Roth ez015890 at bullwinkle.ucdavis.edu
Thu Oct 20 00:18:56 EST 1994


David J. Meyer (meyerdj at biovx1.biology.ucla.edu) wrote:
: In article <01HIDWGZFJEAHTSRCW at JHUVMS.HCF.JHU.EDU>,
: BAI_JN at JHUVMS.HCF.JHU.EDU wrote:

: > Hi, everybody:
: > I deseperately need help for "Single-Step Method of RNA Isolation by Acid
: > Guanidinium Thiocyanate-Phenol-chloroform Extraction". I got this
: protocol from
: > the "RED BOOK" and the original paper-Anal.Biochem 162, P156-159,by
: Chomczynski

: Try adding more CHCl3/isoamyl-OH to the homogenate. This may work if the
: homogenate is not too viscous.


I have used that method for isolating RNA from tissue culture cells and 
have had the same problem.  I have used the original acetate and pH.  The 
addition of more CHCl3 seems to do the trick.  You can actually "see" the 
separation occuring.  I ice for an additional 10 min then spin again when 
the phases don't separate the first time.  By the way, the phases should 
be reasonably separated by the time you go to spin.

Traci Roth Yerby
UC Davis



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